Humoral Immunity: Generation of Antibody Diversity

Question 1

Antibody (Immunoglobulin) - describe structure

Answer 1

2 heavy, 2 light chains (heavy: g, a, m; light: kappa, lambda)
Variable region binds antigen, CH1 support
Constant region- effector functions (activating complement, binding phagocytes)
Disulphide bonds stabilize
VH-CH1-CH2-CH3

Question 2

Somatic recombination vs Differential splicing

Answer 2

Somatic recombination:

V(D)J recombination
Tdt nucleotide addition
Somatic hypermutation
Class switching

Differential splicing:

IgM and IgD
Membrane bound
and secreted Ig

Question 3

Three genetic loci encoding Ig - describe

Answer 3

Three genetic loci encoding Ig

Two for light chain: kappa (κ) and lambda (λ) locus

One for heavy chain

Located on different chromosomes

Question 4

How many antibody genes are inherited?

Answer 4

NONE! No complete genes are inherited, only gene segments

Arranging these gene segments in different combinations generate many Ig sequences

Question 5

Example: VJ recombination of kappa light chain genes (Chromosome 2) - result on AB structure

Answer 5

40 Variable (V) segments

5 Joining (J) segments

Constant region (C) segment

Question 6

Example: VDJ recombination of gamma heavy chain genes (Chromosome 14) - result on AB structure

Answer 6

51 Variable (V) segments

27 Diversity (D) segments

6 Joining (J) segments

Constant region (C) segments

Question 7

Combinatorial diversity of AB - describe

Answer 7

Heavy chain V(51), D(27), J(6) = 8262 (from 94 gene segments inherited)

Light chain: Kappa  V(40), J(5)  = 200 (45 segments inherited)
	       Lambda V(30), J(4) = 120 (30 segments inherited)

     8262 x 200 x 120 = 1.98 x 108

Question 8

Generation of antibody diversity

Answer 8

Multiple germline V, D and J gene segments
Combination V-J and V-D-J joining
Junctional flexibility
P-nucleotide addition
N-nucleotide addition
Combinatorial association of heavy and light chains
Somatic hypermutation during affinity maturation (next lecture)

     1.98 x 108  will become ~3 x1011

Question 9

Recombination signal sequences (RSS) - define and describe function

Answer 9

Recombination signal sequences (RSS) – conserved sequences upstream or downstream of gene segments

‘Turns’ consisting heptamer and nonamer with a 12 or 23 bp spacer

Question 10

Recombination only occurs between what

Answer 10

Recombination only occurs between a segment with a 12bp spacer and a 23bp spacer

Question 11

Junctional flexibility - mechanism = describe

Answer 11

Precise mechanisms unknown

Involves Exonuclease - removes mismatched nucleotides

Question 12

Junctional diversity - describe, +ves/-ves

Answer 12

Junctional flexibility during V(D)J recombination, P and N nucleotide additions

Good: Antibody diversity

Bad: Non-productive rearrangements (incorrect reading frame) – wasteful process

Question 13

Allelic exclusion - describe

Answer 13

Two copies of each Ig gene – one from mother and one from father

In most cases, both genes are expressed

Antibody genes different – Only one heavy chain allele and one light chain allele is expressed

Order of rearrangement: Heavy>kappa>lambda; 1st allele then 2nd

Ensure each B cell makes one type of antibody